1. Field of the Invention
The present invention generally relates to protein-protein interaction, and more particularly, reducing protein-protein interaction between presenilin 2 and a calcium-binding protein to modulate apoptosis and calcium signaling.
2. Background of the Related Art
Alzheimer's disease (AD) is a degenerative disorder characterized clinically by progressive dementia and neuropathologically by the presence of senile plaques and neurofibrillary tangles (NFT). Genetic studies indicate the etiology of AD to be heterogeneous. It is well established that mutation in the genes including, presenilin 1(PS1), and presenilin 2(PS2) genes, cause the majority of early onset of familial Alzheimer's disease (FAD) (i.e., FAD before 65 years of age). Many other genes, including some that act as modifiers or risk factors, appear to be associated with late-onset AD (>65 yr; Corder et al., 1993; Payami et al., 1997; Pericak-Vance et al., 1997; Blacker et al., 1998; Montoya et al., 1998).
Approximately 50% of all FAD cases are linked to the presenilin genes where missense mutations are generally found in residues that are conserved between the two proteins with the rare exceptions of in-frame splice deletions and premature truncations. The mechanisms by which mutations in PS1 and PS2 cause AD are not known, although mutations in these genes appear somehow interconnected as they increase amyloidogenic Aβ fragment accumulation (reviewed by Hardy, 1997).
Human PS1 and PS2 genes are both ubiquitously expressed, but at low protein levels which have lead to difficulties and inconsistencies in their detection and localization. Upon overexpression, the presenilins have been localized to the endoplasmic reticulum (ER) and nuclear envelope (see Kovacs et al., 1996; Janicki and Monteiro, 1997 and references therein) with one group reporting evidence also for cell surface localization (Dewji and Singer, 1997). Endogenous PS1 and PS2 proteins, in turn, have been localized to a variety of structures, including the ER, vesicular structures of the somatodentritic compartment, within axons, at centrosomes and centromeres, and at the plasma membrane (Busciglio et al., 1997; Capell et al., 1997; Li et al., 1997; Ye and Fortini, 1998).
Human PS1 and PS2 proteins are 67% identical, sharing highest similarity in their COOH-terminal sequence and in multiple hydrophobic internal regions that are structurally predicted to form transmembrane domains (TMD). Assuming the presenilins are transmembrane proteins, their topography according to most models is of a protein that weaves through membranes eight times with the NH2— and COOH-terminal domains and the large “loop” spanning the putative sixth and seventh TMD all facing the cytoplasm (see FIG. 1; Hardy, 1997).
Interestingly, presenilins have been implicated in the regulation of programmed cell death (apoptosis). Evidence for such a role was first shown when a cDNA fragment encoding the 103 COOH-terminal amino acids of mouse PS2, termed ALG-3, was isolated in a screen for cDNAs that could rescue T cells from receptor-induced apoptosis (Vito et al., 1996a). This rescue appears to be a consequence of the ALG-3 fragment acting in a dominant negative fashion, since expression of full-length PS2 leads to apoptosis (Vito et al., 1996b). Compared with the apoptosis induced by the overexpression of wild-type PS2 in PC12 and HeLa cells, the FAD PS2-(N141I) mutation causes even higher levels of apoptosis (Deng et al., 1996; Wolozin et al., 1996; Janicki and Monteiro, 1997). Likewise, PS1 overexpression also sensitizes cells to apoptosis (Guo et al., 1996, 1997; Wolozin et al., 1998).
The mechanisms by which presenilins induce apoptosis are not fully understood, but perturbations in calcium Ca2+, oxidative stress (Guo et al., 1996; Keller et al., 1998), destabilization of β-catenin (Zhang et al., 1998), increased signaling by heterotrimeric GTP-binding proteins have been implicated (Wolozin et al., 1996; Smine et al., 1998) and G1 cell cycle arrest (Janicki and Monteiro, 1999; Janicki et al. 2000) have all been implicated.
It seems clear that expressed proteins of the presenilins are somehow built into the apoptotic machinery evidenced by several apoptotic characteristics e.g., cell shrinkage, increased DNA fragmentation, increased Bax expression and caspase activation that have been identified in post-mortem brains of AD patients. Thus, it would be advantages to identify binding partners of the expressed presenilin proteins thereby linking these proteins to known pathways or structures and modulate the activity of the normal and mutant binding partners.